Chemical inhibition of RPA gap protection sensitizes BRCA1-deficient cancers to PARP inhibition

Poly (ADP-ribose) polymerase inhibitors (PARPi) are standard of care for many BRCA1 deficient cancers, though few cures are achieved. We sought to determine if targeting the protection of the single-strand DNA gaps induced by PARPi in BRCA-deficient cancers could increase efficacy. Replication protein A (RPA) participates in critical protein-protein and protein-DNA interactions to protect single-stranded DNA (ssDNA) and support DNA metabolism. We have reported the optimization of small molecule RPA inhibitors (RPAi) that target protein-ssDNA interactions to chemically exhaust RPA and elicit single-agent anticancer activity. RPAi sensitizes cells to PARPi in BRCA1-deficient non-cancerous cells, where ssDNA gap formation drives therapeutic efficacy. We show that RPAi treatment abolishes PARPi-induced replication gap protection, resulting in genomic instability via replication fork degradation and chromosomal integrity, and that, in vivo , the RPA and PARP targeted combination abrogates cancer growth in a BRCA1-mutant breast cancer model. We find that genetic predispositions to ssDNA gap accumulation correlate with RPAi sensitivity, and BRCA1-proficient cells remain sensitive to combination treatment but require more PARP inhibition to increase ssDNA gaps. RPAi-PARPi combination activity in patient-derived ovarian cancer models demonstrates the utility of targeting gap protection to increase PARPi sensitivity and circumvent resistance. Collectively, this work provides a unifying mechanism of chemical RPA exhaustion as a cancer therapeutic strategy.